Repression of E2F1-mediated transcription by the ErbB3 binding protein Ebp1 involves histone deacetylases

Abstract

Ebp1, an ErbB3 binding protein that is a member of the proliferation-associated PA2G4 family, inhibits the proliferation and induces the differentiation of human ErbB positive breast and prostate cancer cell lines. Ebp1 binds the tumor suppressor retinoblastoma protein (Rb) both in vivo and in vitro, and Rb and Ebp1 cooperate to inhibit the transcription of the E2F1-regulated cyclin E promoter. We show here that Ebp1 can inhibit the transcription of other E2F-regulated reporter genes and of several endogenous E2F-regulated genes important in cell cycle progression in both Rb positive and Rb null cells. The Ebp1-mediated transcriptional repression depended on the presence of an E2F1 consensus element in the promoters. A fusion of Ebp1 with the GAL4 DNA binding domain protein had independent transcriptional repression activity that mapped to the C-terminal region of Ebp1. This C-terminal region of Ebp1 bound functional histone deacetylase (HDAC) activity and inhibitors of HDAC significantly reduced Ebp1-mediated repression. Ebp1 bound HDAC2, but not HDAC1, in vitro. An Ebp1 mutant lacking the HDAC binding domain failed to inhibit transcription. Our results suggest that Ebp1 can repress transcription of some E2F-regulated promoters and that one mechanism of Ebp1- mediated transcriptional repression is via its ability to recruit HDAC activity.

Phosphorylation of the ErbB3 binding protein Ebp1 by p21-activated kinase 1 in breast cancer cells

“…Forty-eight hours after transfection, promoter activity was determined using the dual luciferase reporter assay. Wild-type ebp1 inhibited transcription of both the E2F1 and Cyclin D1 genes 50% as reported previously (Zhang et al, 2003). Mutation of T261 to E, mimicking a phosphorylated state, completely abrogated the ability of Ebp1 to repress activity of both the E2F1 and Cyclin D1 promoters ( Figure 5C and D).…”

“…Transcription of both endogenous and exogenous E2F1-regulated genes is repressed by Ebp1 (Zhang et al, 2003). We therefore next investigated the effect of the PAK1 site mutations on the ability of Ebp1 to repress activity of luciferase reporters controlled by the E2F1 and Cyclin D1 promoters.…”

“…ErbB3 binding protein binds ErbB3 in human breast cancer cells Ahn et al, 2006), but dissociates from the receptor in response to HRG . Overexpression of Ebp1 inhibits the transcription of reporter genes controlled by Cyclin D1, Cyclin E and c-myc promoters and the transcription of endogenous E2F1 and c-myc genes via its binding to an E2F1 consensus element (Xia et al, 2001;Zhang et al, 2003;Zhang and Hamburger, 2004). The interaction of Ebp1 with histone deacetylase 2 (HDAC2), Rb and Sin3A is needed for Ebp1 to repress transcription (Zhang et al, 2003Zhang and Hamburger, 2004).…”

“…Overexpression of Ebp1 inhibits the transcription of reporter genes controlled by Cyclin D1, Cyclin E and c-myc promoters and the transcription of endogenous E2F1 and c-myc genes via its binding to an E2F1 consensus element (Xia et al, 2001;Zhang et al, 2003;Zhang and Hamburger, 2004). The interaction of Ebp1 with histone deacetylase 2 (HDAC2), Rb and Sin3A is needed for Ebp1 to repress transcription (Zhang et al, 2003Zhang and Hamburger, 2004). Heregulin, under conditions associated with growth arrest, increases binding of Ebp1 to the E2F1 promoter complex and enhances Ebp1-mediated repression of E2F1-regulated gene transcription (Zhang and Hamburger, 2004).…”

Specificity and heregulin regulation of Ebp1 (ErbB3 binding protein 1) mediated repression of androgen receptor signalling

“…This finding suggests that Ebp1 may play a role in inhibition of AR signalling in the presence of no or extremely low levels of androgens. Ebp1 has the ability to bind DNA (Zhang and Hamburger, 2004) and histone deacetylases (Zhang et al, 2003), and we postulate that Ebp1 might reside on AR regulated promoters in the absence of androgens to inhibit transcription. ChIP assays to demonstrate Ebp1 occupancy on AR promoters are underway in the laboratory.…”